Metalloproteinase activity of cells exposed to the phorbol ester, 12-0-tetradecanoyl-phorbol-13-acetate (TPA), and growth factors was assessed using gelatin sodium dodecyl sulfate (SDS) gel electrophoresis and measurements of type IV collagen degradation. Fibroblasts at different stages of malignant transformation responded variably to TPA. Normal human diploid lung fibroblasts did not respond to TPA at all, but NIH/3T3 cells and transformed fibroblasts expressed up to four times higher type IV collagenolytic activity. Similarly, expression of a 92 kDa gelatinase which is closely associated with the malignant phenotype was increased in the presence of TPA. Conversely, TPA inhibited another major gelatinase of 65 kDa while the 92 kDa gelatinase was induced. The lack of response of normal fibroblasts was not dependent on the stage of confluency in cell culture. Experiments assessing the effect of epidermal growth factor (EFG), transforming growth factor-beta (TGF-beta) and fibroblast growth factor (FGF) on gelatinolytic and type IV collagenolytic activity have been initiated. Preliminary data showed that EGF had no effect on normal and transformed cells, but TGF-beta induced the 92 kDa gelatinase activity of transformed cells in a dose- responsive manner, while the 65 kDa gelatinase activity was only mildly increased. The results obtained so far suggest that a certain degree of cellular atypia, possibly related to the initiation step of the carcinogenic process is necessary for induction of metalloproteinase activity mediated by the tumor promoter TPA, and possibly also by TGF-beta. TPA has opposite effects on the two major gelatinolytic metalloproteinases expressed by malignant cells, i.e., it stimulates the 92 kDa and inhibits the 65 kDa gelatinase.